Iowa State University Capstones, Theses and Retrospective Theses and Dissertations Dissertations 2008 Differential mucosal gene expression modulates the development of murine colitis Zhiping Liu Iowa State University Follow this and additional works at: https://lib.dr.iastate.edu/rtd Part of the Allergy and Immunology Commons, and the Medical Immunology Commons Recommended Citation Liu, Zhiping, "Differential mucosal gene expression modulates the development of murine colitis" (2008). Retrospective Theses and Dissertations. 15866. https://lib.dr.iastate.edu/rtd/15866 This Dissertation is brought to you for free and open access by the Iowa State University Capstones, Theses and Dissertations at Iowa State University Digital Repository. It has been accepted for inclusion in Retrospective Theses and Dissertations by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. Differential mucosal gene expression modulates the development of murine colitis by Zhiping Liu A dissertation submitted to the graduate faculty in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Major: Immunobiology Program of Study Committee: Michael J. Wannemuehler, Major Professor Randy E. Sacco Jesse M. Hostetter Douglas E. Jones Eileen L. Thacker Iowa State University Ames, Iowa 2008 Copyright © Zhiping Liu, 2008. All rights reserved. UMI Number: 3299613 UMI Microform 3299613 Copyright 2008 by ProQuest Information and Learning Company. All rights reserved. This microform edition is protected against unauthorized copying under Title 17, United States Code. ProQuest Information and Learning Company 300 North Zeeb Road P.O. Box 1346 Ann Arbor, MI 48106-1346 ii DEDICATION This dissertation is dedicated to: My daughter Zoe Liu, who was two years old on December 16, 2007; And my son Gavin Liu, who will be born on February 2008; And my wife Jun Jiang. You give purpose and meaning to my life iii TABLE OF CONTENTS Pages TABLE OF CONTENTS iii LIST OF FIGURES v LIST OF ABBREVIATIONS vii CHAPTER 1. GENERAL INTRODUCTION 1 Dissertation organization 1 Literature review 2 Introduction for inflammatory bowel disease(IBD) 2 Development of IBD 13 Mucosal gene expression profiles in IBD 19 Two animal models of intestinal inflammation used in this dissertation 23 References 27 CHAPTER 2. MUCOSAL GENE EXPRESSION PROFILES FOLLOWING THE COLONIZATION OF IMMUNOCOMPETENT GNOTOBIOTIC C3H MICE WITH 46 Helicobacter bilis: A PRELUDE TO COLITIS Abstract 46 Introduction 47 Material and Methods 48 Results 50 Discussion 63 References 66 Supplemental Data 72 CHAPTER 3. EFFECTS OF Helicobacter bilis COLONIZATION ON MURINE 80 COLITIS INDUCED BY DEXTRAN SODIUM SULFATE Abstract 80 Introduction 81 Material and Methods 83 Results 87 Discussion 101 References 105 Supplemental Data 110 CHAPTER 4. ORAL TREATMENT WITH HYPOXOSIDE AMELIORATES 114 Brachyspira hyodysenteriae - INDUCED MURINE COLITIS Abstract 114 Introduction 115 Material and Methods 117 iv TABLE OF CONTENTS (Continued) Pages Results 121 Discussion 130 References 134 Supplemental Data 140 CHAPTER 5. GENERAL CONCLUSION 144 Summary of results 144 Discussion and recommendations for future research 146 ACKNOWLEGEMENTS 161 v LIST OF FIGURES Pages CHAPTER 2 Figure 1 Evaluation of cecal histological lesions of gnotobiotic mice following the 51 colonization of Helicobacter bilis. Figure 2 Scatterplot analysis depicting comparative changes in mRNA expression 53 over time following the colonization of gnotobiotic mice with Helicobacter bilis. Figure 3 Patterns of differentially expressed genes as determined by K-means cluster 54 analysis following the colonization of gnotobiotic mice with Helicobacter bilis. Figure 4 Hierarchical cluster analysis and functional annotation of differentially 55 expressed genes. Figure 5 Analysis of mRNA expression via qRT-PCR in the ceca of gnotobiotic mice 61 following the colonization of Helicobacter bilis. Figure 6 Comparative analysis of mRNA expression of novel colitis-associated 62 genes in the cecal mucosa of gnotobiotic mice after dextran sodium sulfate (DSS) treatment or Brachyspira hyodysenteriae infection. CHAPTER 3 Figure 1 Effect of Helicobacter bilis colonization on the severity of macroscopic and 88 microscopic lesions following exposure to dextran sodium sulfate (DSS). Figure 2 Analysis of mRNA expression of selected genes in the colon of mice treated 93 with DSS alone or treated with H. bilis plus DSS. Figure 3 Analysis of colonic myeloperoxidase (MPO) levels in the colon of mice 94 treated with DSS alone or treated with H. bilis plus DSS. Figure 4 Immunohistochemical analysis of F4/80+ macrophages in colonic mucosa of 96 mice treated with DSS alone or treated with H. bilis plus DSS. Figure 5 Effects of Helicobacter bilis colonization on the total CD3+ T cell and 98 CD19+ B cells in lamina propria and mesenteric lymph node (MLN) following exposure to DSS. Figure 6 Analysis of antigen-specific mesenteric lymph node (MLN) cell 99 proliferation of cell recovered from mice treated with DSS alone or treated with H. bilis plus DSS. Figure 7 Measurement of antibody responses to Helicobacter bilis and altered 100 Schadler’s flora (ASF) in the sera of mice treated with DSS alone or treated with H. bilis + DSS. Figure 8 Analysis of mRNA expression for detoxification-associated genes in the 101 colon of mice treated with DSS alone or treated with H. bilis plus DSS. vi LIST OF FIGURES (Continued) Pages CHAPTER 4 Figure 1 The effects of hypoxoside treatment on gross lesion and weight change 122 induced by Brachyspira hyodysenteriae infection in mice. Figure 2 The effects of hypoxoside treatment on the cecal histological lesions 123 induced by Brachyspira hyodysenteriae infection in mice. Figure 3 The effects of hypoxoside treatment on cecal myeloperoxidase (MPO) 124 activity in mice. Figure 4 The effects of hypoxoside treatment on gene expression profiles in NF-қB 125 pathway. Figure 5 The effects of hypoxoside treatment on active cecal mucosal p65 protein 128 levels in mice. Figure 6 The effects of hypoxoside treatment on cecal epithelial cell proliferation 129 shown by BrdU staining. Figure 7 The effects of hypoxoside treatment on the number of cecal Brachyspira 131 hyodysenteriae. vii LIST OF ABBREVIATION Abbreviation Full names ANOVA Analysis of variance AP-1 Activator protein-1 APC Antigen presenting cell ASF Altered Schaedler's flora B. hyodysenteriae Brachyspira hyodysenteriae B3galt5 UDP-Gal:betaGlcNAc beta 1,3-galactosyltransferase, polypeptide 5 Bcl3 B-cell leukemia/lymphoma 3 BrdU 5-bromo-2-deoxyuridine CAR Constitutive androstane receptor (Nr1i3) (Nuclear receptor subfamily 1, group I, member 3 ) Ccl5 Chemokine (C-C motif) ligand 5 Ccl8 Chemokine (C-C motif) ligand 8 Ccr10 Chemokine (C-C motif) receptor 10 Ccr5 Chemokine (C-C motif) receptor 5 CD Crohn's disease cDNA Complementary DNA CDT Cytolethal distending toxin Ceacam12 CEA-related cell adhesion molecule 12 Ces1 Carboxylesterase 1 Ces3 Carboxylesterase 3 Chuk(IKKα) Conserved helix-loop-helix ubiquitous kinase (I-kappa-B kinase-alpha) Cyp4b1 Cytochrome P450, family 4, subfamily b, polypeptide 1 DAVID The Database for Annotation, Visualization and Integrated Discovery DC Dendritic cells DPI Days post infection DSS Dextran sodium sulfate EHS Enterohepatic Helicobacter species ELISA Enzyme-Linked ImmunoSorbent Assay FDR False discovery rate Fos FBJ osteosarcoma oncogene Foxp3 Forkhead box P3 Fut2 Fucosyltransferase 2 GAPDH Glyceraldehyde-3-phosphate dehydrogenase GBP Guanylate nucleotide binding protein GO term Gene-ontology term Gzmb Granzyme B H&E Hematoxylin and eosin stain H. bilis Helicobacter bilis HLAB27 Human Leukocyte Antigen B27 IBD Inflammatory bowel disease Ifi47 Interferon gamma inducible protein 47 viii LIST OF ABBREVIATION (Continued) Abbreviation Full names IFNγ Interferon gamma Map3K14 mitogen-activated protein kinase kinase kinase 14 (NIK) (NF kappa B-inducing kinase) IL Interleukin Irf4 Interferon regulatory factor 4 ItgaL(Cd11a) Itegrin alpha L (CD11a molecule) Itgam (Cd11b) Integrin alpha M (CD11b molecule) Itgb2(Cd18) Integrin, beta 2 (CD18 molecule) Lcp Lymphocyte cytosolic protein 1 LFA-1 Lymphocyte function-associated antigen 1 LPL Lamina propria lymphocytes Ltbr Lymphotoxin B receptor Mcpt Mast cell protease Mdr1a Multi-drug resistance 1a MLN Mesenteric lymph nodes MPO Myeloperoxidase Muc2 Mucin 2 NFκb Nuclear factor kappa B NFκb1(p50) Nuclear factor of kappa light chain gene enhancer in B-cells 1, p105 (p50) NFκb2(p52) Nuclear factor of kappa light polypeptide gene enhancer in B-cells 2, p49/p100 (p52) Nod2 Nucleotide-binding oligomerization domain containing 2 p65 p65 NF-kappa B (RelA) ( v-rel reticuloendotheliosis viral oncogene homolog A ) Pparγ Peroxisome proliferator-activated receptor gamma PXR Pregnane x receptor qPCR Quantitative polymerase chain reaction Reg Regenerating islet-derived RT-PCR Reverse transcriptase-polymerase chain reaction SCID Severe combined immunodeficiency SEM Standard error of measurement Socs3 Suppressor of cytokine signaling 3 TCR T cell receptor TGFβ Transforming growth factor, beta TLR Toll-like receptor TNBS Trinitrobenzesulfonic acid TNF Tumor necrosis factor Tnfsf13b Tumor necrosis factor (ligand) superfamily, member 13b (BAFF) (B-cell activating factor) Treg Regulatory T cells UC Ulcerative colitis Ugt8a UDP galactosyltransferase 8A 1 CHAPTER 1. GENERAL INTRODUCTION General Introduction and Dissertation Organization Inflammatory bowel diseases (IBD), including Crohn’s disease